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2024-12-16
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Bacteria and Yeast Strain Development: Applications and Recent Advances

December 16, 2024
1:17 pm

Bacteria and Yeast Strain Development: Applications and Recent Advances

By Dr. Pierre-Henri Ferdinand

Head of Product

From producing lifesaving therapeutics like insulin to creating eco-friendly biofuels, engineered strains of bacteria and yeast offer solutions to pressing global challenges. Advances in synthetic biology and automation are driving this field forward, enabling efficient processes and novel applications. However, challenges like single clone isolation, growth condition optimization, and regulatory hurdles persist. This blog explores the applications of bacteria and yeast strain development, highlighting challenges and advances in this rapidly evolving field.

Applications of Strain Development

The development of bacterial and yeast strains has diverse applications. These microorganisms are utilized to produce therapeutic products and as integral components in various industrial processes.

Therapy Production

Bacteria and yeast can be modified to produce therapeutic proteins. Early applications included producing human insulin in E. coli to treat diabetes. However, more recent applications include producing erythropoietin (EPO), human growth hormone, cytokines, and recombinant proteins for use as antigens in vaccines. While mammalian cells have overtaken bacterial cells in producing many protein-based therapies, there are still applications where bacteria are preferred, such as those with less post-translational complexity (Rettenbacher et al., 2022). These include interferons (e.g., IFNα-2b), cytokines (e.g., G-CSF, TNFs), hormones (e.g., insulin glargine, hGH), and interleukins (e.g., IL-2, IL-11), which are commonly produced in E. coli and yeasts (Rettenbacher et al., 2022).

Synthetic Biology

The core principle of synthetic biology is to modify biological systems and organisms to perform specific functions. Emerging applications include the generation of oncolytic bacteria to treat cancer and the creation of living biosensors to detect and respond to disease and pollution biomarkers in real time (Kiaheyrati et al., 2024; Wan et al., 2021).

Agriculture and Biofuels

Biofuels offer an important environmentally friendly alternative to fossil fuels. They are produced by engineered yeast cell lines metabolizing different organic materials (called biomass), including plants, algae, and waste (Keasling et al., 2021). Bacterial cells are important as biopesticides to protect economically important crops from pests. The development of these lines comes as the industry moves towards more environmentally friendly alternatives to conventional chemical pesticides (Ayilara et al., 2023; Ragasruthi et al., 2024).

Challenges in Strain Development

Despite the promise of bacteria and yeast strain development for many applications, there remain several technical challenges that hinder their full potential.

Single Clone Isolation

Isolating single clones from transfection pools and complex mixtures is a major challenge in bacteria strain development. This poses a significant obstacle to workflows, as ensuring clonality is crucial to prevent contamination and enables more efficient, high-throughput clone screening.

Growth Conditions

Diverse bacterial and yeast strains require different conditions for optimal growth. This is particularly important when scaling up bacterial and yeast cells after successful isolation. Finding optimized growing conditions can be challenging because there are a huge number of parameters that need to be considered, including oxygen concentration, growth substrate, pH, and other factors. These parameters can fluctuate in large-scale cultures, negatively impacting production (Xu et al., 2024).

The S.NEST from CYTENA provides continuous monitoring of dissolved oxygen and pH to ensure that cells are cultured in optimal conditions
(Fig. 1).

Figure 1. The S.NEST from CYTENA allows researchers to incubate up to four 24- or 96-well plates simultaneously to boost laboratory efficiency.

Regulatory Considerations

The use of genetically modified organisms as therapeutic agents or in the production of therapeutic agents is tightly controlled by regulatory bodies. Researchers must be vigilant to ensure their cell lines are free from contaminants and remain genetically stable and fit for purpose throughout the production and regulatory process.

Advances in Imaging

Advances in microfluidics and imaging technology have facilitated more streamlined and accurate isolation of single cells.

Seeding

Advanced imaging technologies such as those found in the B.SIGHT from CYTENA have made it simple to detect the dispensing of single cells into multiwell plates. Within the B.SIGHT instrument, this enables single-cell omics without requiring cultivation and integrates seamlessly into sequencing workflows. The B.SIGHT uses dual imaging and extremely high-resolution optics to ensure that even the smallest cells, whether labeled or unlabelled, can be detected as they are dispensed into wells (Fig. 2).

Figure 2. The B.SIGHT enables rapid seeding of both labeled and non-labeled microbial cells with advanced imaging to provide robust evidence of clonality.

Monitoring and Analysis

Advanced image analyses allow for high throughput phenotypic characterization and screening of huge numbers of individual microbial clones. This facilitates the selection of optimal clones based on various characteristics, such as growth rate and response to stimuli (Zahir et al., 2019). Pooled screening is a method that uses high-content imaging coupled with genotyping to identify the phenotypic changes induced by modifications to different genes (Feldman et al., 2022). This helps researchers to rapidly link specific genetic perturbations to observable traits, enabling functional genomics studies, the identification of gene functions, and the discovery of potential targets for therapeutic or industrial applications.

Automation and Future Directions

Automated dispensing of bacterial and yeast cells means that single cells can be rapidly seeded into multiwell plate formats from complex mixtures and transfection pools. Automation removes significant sources of error, such as contamination, which can severely compromise strain development. When coupled with advances in microfluidics, automation allows for the rapid dispensing of tiny volumes containing individual cells. Scaling down isolation and culturing processes significantly reduces the cost of development workflows and enables higher throughput in screening processes.

Future advances in this area include the culturing of microbial consortia, which involves cultivating multiple species in tandem to produce metabolites and other products, which could enhance efficiency, reduce production costs, and enable the synthesis of complex compounds that are challenging to produce with single-species systems (Mittermeier et al., 2023). Advancements in bacterial cell systems are underway to better emulate the protein-folding mechanisms of mammalian cells, potentially enabling significant improvements in their capacity to produce therapeutic-grade proteins in the future (Jayakrishnan et al., 2024).

Conclusion

Despite challenges in isolation, scalability, and regulatory requirements, advancements in imaging, automation, and synthetic biology promise to accelerate progress in bacteria and yeast strain development. Future directions, including culturing microbial consortia and generating microbes more suitable for mammalian protein production, indicate a thriving field poised to address global needs. As these technologies mature, they offer a pathway to sustainable, efficient, and transformative solutions in healthcare, agriculture, and beyond.

The B.SIGHT is revolutionizing how researchers isolate single microbial cells for various downstream applications. Talk to a CYTENA expert today to learn more about this exciting piece of technology.

References

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Feldman, D., Funk, L., Le, A., Carlson, R. J., Leiken, M. D., Tsai, F., Soong, B., Singh, A., & Blainey, P. C. (2022). Pooled genetic perturbation screens with image-based phenotypes. Nature Protocols, 17(2), 476–512.
Jayakrishnan, A., Wan Rosli, W. R., Tahir, A. R. M., Razak, F. S. A., Kee, P. E., Ng, H. S., Chew, Y.-L., Lee, S.-K., Ramasamy, M., Tan, C. S., & Liew, K. B. (2024). Evolving Paradigms of Recombinant Protein Production in Pharmaceutical Industry: A Rigorous Review. Sci, 6(1), 9.
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